Intestinal epithelial culture under an air-liquid interface: a tool for studying human and mouse esophagi.

This study investigated whether an intestinal epithelial culture method can be applied to mouse and human esophageal cultures. The esophagi harvested from 1-day-old mice and adult humans were maintained in collagen gels. A commercially available culture medium for human embryonic stem cells was used for the human esophageal culture. We discovered that the intestinal epithelial culture method can be successfully applied to both mouse and human esophageal cultures. The long-term cultured esophageal organoids were rod-like luminal structures lined with myofibroblasts. We discovered that regeneration of the esophageal mucosal surface can be almost completely achieved in vitro, and the advantage of this method is that organoid cultures may be generated using host-derived fibroblasts as a niche. This method is a promising tool for mouse and human research in intestinal biology, carcinogenesis, and regenerative medicine.

Circulating tumour cell-derived plastin3 is a novel marker for predicting long-term prognosis in patients with breast cancer.

BACKGROUND: Identification of promising biomarkers that predict the prognosis of patients with breast cancer is needed. In this study, we hypothesised that the expression of the epithelial-mesenchymal transition-related biomarker plastin3 (PLS3) in peripheral blood could be a prognostic factor in breast cancer. METHODS: We examined PLS3 expression in breast cancer cell lines with epithelial and mesenchymal traits and in circulating tumour cells (CTCs) obtained from the peripheral blood of breast cancer patients. We investigated PLS3 expression in the peripheral blood of 594 patients with breast cancer to evaluate the clinical significance of PLS3 expression. RESULTS: Robust PLS3 expression was observed in different breast cancer cell lines (Hs578t, MCF-7, MDA-MB-468, and MDA-MB-231) as well as in a bone marrow derived cancer cell line (BC-M1). In both the training (n=298) and validation (n=296) sets, PLS3 expression was observed in CTCs of patients with breast cancer. PLS3-positive patients showed significantly poorer overall and disease-free survival than PLS3-negative patients (P=0.0001 and 0.003, respectively). Subset analysis revealed that this prognostic biomarker was relevant in patients with stage I-III cancer, particularly in patients with luminal-type and triple-negative-type tumours. CONCLUSIONS: These data demonstrated that PLS3 was expressed in CTCs undergoing the epithelial-mesenchymal transition in patients with breast cancer. Furthermore, PLS3 may be an excellent biomarker for identifying groups at risk of recurrence or with a poor prognosis.

Biological significance of fluorine-18-α-methyltyrosine (FAMT) uptake on PET in patients with oesophageal cancer.

PURPOSE: (18)F-FAMT as an amino-acid tracer for positron emission tomography (PET) is useful for detecting human neoplasms. (18)F-FAMT is accumulated in tumour cells solely via L-type amino-acid transporter 1 (LAT1). This study was conducted to investigate the biological significance of (18)F-FAMT uptake in patients with oesophageal cancer. METHODS: From April 2008 to December 2011, 42 patients with oesophageal cancer underwent both (18)F-FAMT PET/CT and (18)F-FDG PET/CT before surgical treatment. The immunohistochemical analysis of LAT1, CD98, Ki-67, CD34, p53, p-Akt and p-mTOR was performed on the primary lesions. In vitro experiments were performed to examine the mechanism of (18)F-FAMT uptake. RESULTS: High uptake of (18)F-FAMT was significantly associated with advanced stage, lymph node metastasis and the expression of LAT1, CD98, Ki-67 and CD34. LAT1 expression yielded a statistically significant correlation with CD98 expression, cell proliferation, angiogenesis and glucose metabolism. In vitro experiments revealed that (18)F-FAMT was specifically transported by LAT1. CONCLUSIONS: The uptake of (18)F-FAMT within tumour cells is determined by the LAT1 expression and correlated with cell proliferation and angiogenesis in oesophageal cancer. The present experiments also confirmed the presence of LAT1 as an underlying mechanism of (18)F-FAMT accumulation.

PICT1 regulates TP53 via RPL11 and is involved in gastric cancer progression.

BACKGROUND: The TP53 pathway is frequently inactivated in human cancers. PICT1 (also known as GLTSCR2) is a novel regulator of the MDM2-TP53 pathway via its interaction with the ribosomal protein RPL11 in the nucleolus. However, the clinical significance of PICT1 in gastric cancer remains unknown. METHODS: To evaluate PICT1 function, we used shRNA to inhibit PICT1 expression in gastric cancer cells that expressed wild-type TP53. PICT1 expression and TP53 mutation status were quantified in 110 cases of primary gastric cancer to explore the impact of PICT1 expression levels on gastric cancer. RESULTS: Deficiency of PICT1 significantly impaired cell proliferation and colony formation via TP53-mediated cell cycle arrest. Following induction of PICT1 deficiency, RPL11 translocated out of the nucleolus. Of the 110 gastric cancer samples tested, 70 (63.6%) and 40 (36.4%) tumours expressed wild-type and mutant TP53, respectively. In gastric cancer patients with wild-type TP53 tumours, patients with relatively low PICT1 expression levels had a better prognosis compared with high expression level patients (P=0.046). CONCLUSION: The findings suggest that PICT1 has a crucial role in gastric cancer progression by regulating the MDM2-TP53 pathway through RPL11. Clinically, PICT1 expression is a novel prognostic parameter in gastric cancer patients with wild-type TP53 tumours.

Emergence of highly degenerate excited states in the frustrated magnet MgCr2O4.

High degeneracy in ground states leads to the generation of exotic zero-energy modes, a representative example of which is the formation of molecular spin-liquid-like fluctuations in a frustrated magnet. Here we present single-crystal inelastic neutron scattering results for the frustrated magnet MgCr(2)O(4), which show that a common set of finite-energy molecular spin excitation modes is sustained in both the liquid-like phase above magnetic ordering temperature T(N) and an ordered phase with an extremely complex magnetic structure below T(N). Based on this finding, we propose the concept of high degeneracy in excited states, which promotes local resonant elementary excitations. This concept is expected to have ramifications on our understanding of excitations in many complex systems, including not only spin but also atomic liquids, complex order systems, and amorphous systems.

Bone marrow and peripheral blood expression of ID1 in human gastric carcinoma patients is a bona fide indicator of lymph node and peritoneal metastasis.

Recent studies have showed that the bone marrow-derived endothelial progenitor cells play critical roles in metastasis and that ID1 is required in metastasis as regulator of angiogenesis. Therefore, we investigated the clinical significance of ID1 mRNA expression in bone marrow and peripheral samples in patients with gastric cancer. Two hundred and eighty-nine bone marrow and 196 peripheral blood samples from gastric cancer patients were collected and analysed by quantitative RT-PCR for ID1. The ID1 protein expression in one bone marrow, three metastatic lymph nodes and three peritoneal disseminated tumours was examined by immunohistochemical methods. In both bone marrow and peripheral blood samples, ID1 mRNA expression in the metastatic group was significantly higher than in any other group (P=0.003, P=0.0001, respectively) and significantly associated with lymph node metastasis and peritoneal dissemination. The cells in bone marrow with metastatic cancer stained strongly with ID1 compared with those of healthy volunteers. The expression of ID1 mRNA in bone marrow and peripheral blood was significantly associated with lymph node metastasis and peritoneal dissemination, and therefore constitutes a predictable marker for lymph node metastasis and peritoneal dissemination.

Myoferlin regulates cellular lipid metabolism and promotes metastases in triple-negative breast cancer.

Myoferlin is a multiple C2-domain-containing protein that regulates membrane repair, tyrosine kinase receptor function and endocytosis in myoblasts and endothelial cells. Recently it has been reported as overexpressed in several cancers and shown to contribute to proliferation, migration and invasion of cancer cells. We have previously demonstrated that myoferlin regulates epidermal growth factor receptor activity in breast cancer. In the current study, we report a consistent overexpression of myoferlin in triple-negative breast cancer cells (TNBC) over cells originating from other breast cancer subtypes. Using a combination of proteomics, metabolomics and electron microscopy, we demonstrate that myoferlin depletion results in marked alteration of endosomal system and metabolism. Mechanistically, myoferlin depletion caused impaired vesicle traffic that led to a misbalance of saturated/unsaturated fatty acids. This provoked mitochondrial dysfunction in TNBC cells. As a consequence of the major metabolic stress, TNBC cells rapidly triggered AMP activated protein kinase-mediated metabolic reprogramming to glycolysis. This reduced their ability to balance between oxidative phosphorylation and glycolysis, rendering TNBC cells metabolically inflexible, and more sensitive to metabolic drug targeting in vitro. In line with this, our in vivo findings demonstrated a significantly reduced capacity of myoferlin-deficient TNBC cells to metastasise to lungs. The significance of this observation was further supported by clinical data, showing that TNBC patients whose tumors overexpress myoferlin have worst distant metastasis-free and overall survivals. This novel insight into myoferlin function establishes an important link between vesicle traffic, cancer metabolism and progression, offering new diagnostic and therapeutic concepts to develop treatments for TNBC patients.

Proteoglycans synthesized in calluses at various stages of fracture healing in rats.

The sequential cartilage and bone formation was observed in fracture calluses which were formed at fibula diaphysis of Sprague-Dawley rats. The proteoglycans synthesized in the calluses at different stages during fracture healing were labeled in vitro with [35S]sulfate and then analyzed by sucrose density gradient centrifugation. A heavy proteoglycan, in which chondroitin 4-sulfate accounted for approx. 90% of total radioactivity, was predominantly synthesized in addition to light, dermatan sulfate-containing proteoglycans in day-10 and day-20 calluses, when cartilaginous areas were predominant in these calluses. The synthesis of the heavy proteoglycan started around day 7, when chondrogenesis started locally in the callus, and ceased by day 30, when cartilage had been replaced by newly formed bone. The heavy proteoglycan had chemical and physical properties similar to those of the major proteoglycan synthesized by bone matrix-induced cartilages, but different from those of the major one synthesized by the epiphyseal cartilage of neonatal rats. These findings suggest that the sequential molecular transitions observed in fracture healing differ from those in the endochondral ossification of embryonic skeletal tissues but resemble those in bone matrix-induced bone formation.

What are biomechanics and biomechanical behaviour?

So-called bioengineering is, indeed, of an interdisciplinary nature. It covers very huge fields and concerns many various concepts and ideas. Bioengineering, in this sense, appears to include the fields, in terms of terminology, of bioscience, biotechnology, biomaterials, biomechanical behaviour, biomechanics, and so forth. On the other hand, it seems to be difficult to draw distinctions among these fields, as is often the case in such interdisciplinary science and technology. Then the question may arise what are bioengineering, biotechnology, biomaterials, biomechanical behaviour, biomechanics, etc.? Herein, from this point of view, an attempt will be made to consider the interaction among the concepts and the prevalent approaches in each separate field called biomaterials, biomechanical behaviour, and biomechanics. Also, some approaches and methodologies that remain unknown have been pointed out for future new development of the research in the mechanical behaviour of bio- and biomedical materials and in bioengineering.

Some philosophies for standardizations of biomaterials and related devices.

From the viewpoint of the target of biomaterials, the needs for biomedical reliability as well as development of novel function is discussed. Some approaches to guidelines or standardization of biomaterials and the related devices are pointed out.

The mechanical test method of cardiovascular and related biomaterials.

Mechanical testing methods were developed by us using the pulsatile internal pressure through-flow system. The methods were applied to estimate the permeability and fatigue strength of a vascular substitute, the mechanical behaviour of anastomosed site between vascular substitute and blood vessel, and the mechanical behaviour of the blood vessel itself. A testing method is also proposed for puncturing a vascular substitute which is used as blood access for hemodialysis. The mechanical behaviour of punctured vascular substitute and the puncturing technique itself are investigated. The fatigue-creep testing method of cellophane membrane is also reported to estimate the durability of cellophane membrane by using the circulating pulsatile internal pressure through-flow system. The mechanical testing method for visco-elastic characteristic of biomaterials is discussed.

Effects of pulsatile flow pattern and each stage of pressure increasing, constant and decreasing, respectively upon inelastic deformation of blood vessel.

The aim of this study is to elucidate the relation of the inelastic expansive deformation of natural blood vessel and the degradation of elasticity to the time pattern of circulating pulsatile pressure flow. When pulsatile pressure amplitude (delta P = Pmax - Pmin) becomes smaller due to Pmin increasing, the blood vessel is subject to creep deformation. In this sense, pulsating pressure will play a role in avoiding creep effect. Fluctuation of maximum pressure Pmax will induce the increase of inelastic deformation and the decrease of rigidity of the blood vessel. The inelastic deformation and the decrease of rigidity in blood vessel is induced at the stage of pressure amplitude rising from a lower one to a higher, but not during pressure amplitude (delta P) kept constant nor at the pressure amplitude decreasing stage. In order to reduce the degradation of mechanical properties of blood vessel, it may be effective to avoid the increase of Pmin and the variability of Pmax.

Microscopical imaging of hydroxyapatite/mica composite and packed hydroxyapatite structure--an atomic force microscopy investigation.

A fine grained hydroxyapatite/mica composite material was studied by the atomic force microscopy method and the results were compared with results of atomic force microscopy studies of very five grained hydroxyapatite. In the investigation it was found that the fractal dimension diagram from the atomic force microscopy studies is a tool by which mechanical properties on the surface of the material can be predicted. The two investigated materials were found to show self-similarity properties, i.e., they are identical on the surface. The information given by the fractal dimension is important, and the fractal dimension analysis is an important tool in future designing and engineering of, especially, bioceramics and composites.

Fracture toughness of hydroxyapatite/mica composite, packed hydroxyapatite, alumina ceramics, silicon nitride and -carbide.

By using the fracture toughness estimation method based on two-dimensional map, it was found that the ductility of the high porosity hydroxyapatite/mice composite was comparable with silicon carbide. It was measured to be higher than that of packed hydroxyapatite. Alumina ceramics with more than 96% aluminium oxide showed a higher fracture toughness than the composite material. When bending strength was compared, the strength of the composite was two or three times lower than that of packed hydroxyapatite and much lower than the other studied materials. The composite material showed high porosity, which in turn gives it a lower bending strength. However, the high porosity is more favourable for biocompatibility.

Osteogenic differentiation of cultured marrow stromal stem cells on surface of microporous hydroxyapatite based mica composite and macroporous synthetic hydroxyapatite.

In order to investigate the significance of hydroxyapatite based microporous composite (HA/mica composite) surfaces and a macroporous synthetic hydroxyapatite, rat marrow cell culture, which shows osteogenic differentiation, was carried out on six different culture substrata (two control culture dishes, two identical HA/mica composites, and two identical macroporous synthetic hydroxyapatites). A culture period of two weeks in the presence of beta-glycerophosphate (BGP), ascorbic acid, and dexamethasone resulted in abundant mineralized nodule formations that were positive for alkaline phosphatase (ALP) stain. The stain on the macroporous synthetic hydroxyapatite and the HA/mica composites were intense, the enzyme activity being about double that of control culture dishes. These data indicate that the synthetic macroporous hydroxyapatite surface and the HA/mica composite surface promotes osteoblastic differentiation.

Hybrid concept on the mechanical test method of small caliber blood vessel.

A mechanical test on small caliber blood vessel is difficult because the strength is very low and the dimension is very small. In the present paper a multifunctional axial test apparatus design is proposed. This has the characteristics such that the load cell and the environmental container for the specimen are built up in terms of cassette, respectively, and thus both are easy to equip and take off, that is, easy to replace. Also, the test methodology by internal pressure for small caliber blood vessels has been proposed. By using both of these methodologies, the characteristics of the mechanical behavior of small caliber blood vessels, such as compliance, stress relaxation, and viscoelastic property have been clarified. An attempt has been made on the application of this method to a clinical case.

Acoustical imaging and processing of blood vessel and the related materials using ultrasound Doppler effect.

In the present paper a method is proposed to measure the degree of the degradation of the elasticity in natural blood vessel and the related materials by using ultrasound Doppler effect. It was found that the deformation rate and its acceleration in the radial direction of the blood vessel can be detected by acoustical imaging and processing using this method. These results were proven to correspond to the degree of the degradation of the elasticity, that is, the degree of viscoelasticity in the blood vessel from the wave versus time pattern detected and its simple analysis. This method was applied to predicting the arteriosclerosis of blood vessels of humans by acoustical imaging and processing uninvadedly, as the characteristics of viscoelasticity in blood vessels.

Algorithm of the noninvasive diagnosis method on the atherosclerosis by ultrasonic Doppler effect.

Previously we proposed Acoustical Imaging and Processing Method to measure the viscoelastic property of the blood vessels of a canine and the related materials using Ultrasonic Doppler Effect Measurement. Furthermore, its theoretical foundation was presented. In this paper, this method is applied to measure the viscoelastic mechanical property, that is, the mechanical degradation of human blood vessels by a percutaneous noninvasive method. Based on these results, we proposed the algorithm of the noninvasive estimation method on the viscoelastic mechanical property of the blood vessel by using Doppler Effect Sensor. This method makes it possible to discriminate the blood vessel with arteriosclerosis from a normal blood vessel. Clinical applications were successfully performed using our developed computer software based on our proposed algorithm.

[Puncture technology and selective renal biopsy].

An experimental study of puncture technology for the kidney: We investigated the strength characteristics of tissues which are penetrated by the puncture needle for the kidney using a tension test. The tensile strength of the fascia, the muscle, the renal capsule, the renal parenchyma and the renal pelvis were 13.9, 1.1, 29.5, 0.5 and 21.4 kg/cm2, respectively. As the strengths differ, the needles for each tissue clearly need to be changed. Needle tip shape and needle material for kidney puncture were investigated by compression test, acryl resin injection into the renal vasculature, stereo-microscopy and scanning electron microscopy. These investigations revealed that the most suitable needle tip was a sharp cone shape, and material with a smooth surface and some flexibility such as Derlin seemed to be the most appropriate for the puncture needle for the kidney. Using the acryl resin injection method, stereo-microscopy and scanning electron microscopy, we studied the relative safety of single stage puncture and the repeated dilatation method to establish percutaneous nephrostomy. Single stage puncture seems to cause less damage to the kidney than the repeated dilatation method. Selective renal biopsy: A new puncture system using real-time ultrasound was developed at our clinic in 1978. The puncture procedure is monitored in 2 dimensional real-time images by a mechanical sector scanner equipped with an attachment for needle guidance. The application of the system to percutaneous renal biopsy is called "selective renal biopsy" because the specimen can be obtained selectively from any portion of the kidney.(ABSTRACT TRUNCATED AT 250 WORDS)

[A study of the simulation model of the lower urinary tract for urodynamics--(the first report)--theoretical evaluation of hydrodynamic model].

From the view point of urodynamics, the lower urinary tract might be able to be simulated by a hydrodynamic model, consisting of a spherical balloon corresponding to the bladder and a circular tube to the urethra. In this hydrodynamic model, the diameter of the tube corresponding to the posterior urethra was assumed to become smaller with increase of the prostatic pressure according to the development of BPH. On this assumption, Bernoulli's equation might be adopted. Because of the turbulent flow in the tube, the velocity of flow was expressed as a function of the invariants, which indicated the pressure in the balloon, the respective lengths and diameters of narrow tube and wide tube, coefficients due to pipe friction, pipe fitting and pipe enlargement. The findings suggested that the smaller the diameter of the narrow tube, the smaller the flow volume showed with the development of BPH. In conclusion the urodynamics in the lower urinary tract could be simulated quantitatively by this model. The study may lead to the development of the urodynamic simulation model for the lower urinary tract.